犬弓形虫巢式PCR检测方法的建立及初步应用  

Development and Application of Nested PCR Assay for Detection of Toxoplasma gondii Infection in Dogs

作　　者：柳方远 李双星 印春生[1] 吉婧 朱明哲 刘业兵[1] LIU Fangyuan;LI Shuangxing;YIN Chunsheng;JI Jing;ZHU Mingzhe;LIU Yebing(China Institute of Veterinary Drug Control,Beijing 10081,China;Animal Husbandry and Veterinary,Zhangjiagang 215600,China;Qingdao Agricultural University,Qingdao 266109,China)

机构地区：[1]中国兽医药品监察所,北京100081 [2]张家港市畜牧兽医站,张家港215600 [3]青岛农业大学,青岛266109

出　　处：《中国动物传染病学报》2024年第1期129-134,共6页Chinese Journal of Animal Infectious Diseases

基　　金：国家重点研发计划项目(2016YFD0501004)。

摘　　要：弓形虫病是一种重要的人兽共患病,犬是弓形虫重要的中间宿主,有必要建立一种快速、灵敏的犬弓形虫病检测方法。根据GenBank中弓形虫529 bp基因重复序列设计巢式PCR引物,通过优化反应条件建立巢式PCR检测方法。特异性试验结果显示,该方法对牛巴贝斯虫、环形泰勒虫、犬新孢子虫的基因扩增无条带;灵敏度试验结果显示,该方法最低可以检出0.134 pg的弓形虫基因,比目前国标PCR方法高100倍。对感染弓形虫犬的组织样品检测发现,巢式PCR能在感染犬的不同组织中检测出弓形虫基因。建立的犬弓形虫巢式PCR检测方法特异性强、灵敏度高、重复性好,为该病的检测奠定了基础。Toxoplasmosis is a global zoonosis,which requires a rapid and sensitive method for the detection of canine infection.Nested PCR primers were designed according to the repetitive 529 bp DNA fragment of Toxoplasmagondii in GenBank.The nested PCR detection method was then developed and optimized for its reaction conditions.The results of specificity test showed that this method had no amplification of Cattle Babesia,TheileriaannulataandNeospora.The sensitivity test results showed that it was 100 times higher than the national standard PCR and the method detected the lowest T.gondii gene of 0.134 pg.Testing of tissue samples from dogs infected withT.gondiishowed good detection by this nested PCR.Taken together,the nested PCR method developed here had strong specificity,high sensitivity and good repeatability,which provided technical support for the detection of T.gondii infection and prevention and control of the disease.

关 键 词：弓形虫 巢式PCR 犬 

分 类 号：S854.4[农业科学—临床兽医学]